Two novel oligosaccharides formed by 1F-fructosyltransferase purified from roots of asparagus (Asparagus officinalis L.)

Two novel oligosaccharides, tetra-and penta-saccharides were synthesized by fructosyl transfer from 1-kestose to 4G-beta-D-galactopyranosylsucrose with a purified 1F-fructosyltransferase of asparagus roots and identified as 1F-beta-D-fructofuranosyl-4G-beta-D-galactopyranosylsucrose, O-beta-D-fructofuranosyl-(2-->1)-beta-D-fructofuranosyl-O-[beta-D-galactopyranosyl-(1-->4)]-alpha-D-glucopyranoside and 1F(1-beta-D-fructofuranosyl)2-4G-beta-D-galactopyranosylsucrose, [O-beta-D-fructofuranosyl-(2-->1)]2-beta-D-fructofuranosyl-O-[beta-D-galactopyranosyl-(1-->4)]-alpha-D-glucopyranoside, respectively. Both oligosaccharides were scarcely hydrolyzed by carbohydrase from rat small intestine. Human intestinal bacterial growth by 1F-beta-D-fructofuranosyl-4G-beta-D-galactopyranosylsucrose was compared with that by the tetrasaccharides, stachyose and nystose. Bifidobacteria utilized 1F-beta-D-fructofuranosyl-4G-beta-D-galactopyranosylsucrose to the same extent as stachyose or nystose. On the other hand, the unfavorable bacteria, Clostridium perfringens, Escherichia coli and Enterococcusfaecalis, that produce mutagenic substances did not use the synthetic oligosaccharide.     

Synthesis of two phosphate-containing "heptasaccharide" fragments of the capsular polysaccharides of Streptococcus pneumoniae types 6A and 6B.

The "heptasaccharides" O-alpha-D-galactopyranosyl-(1----3)- O-alpha-D-glucopyranosyl-(1----3)-alpha, beta-L-rhamnopyranose 2'-[O-alpha-D-galactopyranosyl-(1----3)-O-alpha-D-glucopyranosyl- (1----3)-O-alpha-L-rhamnopyranosyl-(1----3)-D-ribit-5-yl sodium phosphate] (25) and O-alpha-D-galactopyranosyl- (1----3)-O-alpha-D-glucopyranosyl-(1----3)-alpha, beta-L-rhamnopyranose 2'-[O-alpha-D-galactopyranosyl-(1----3)-O-alpha-D-glucopyranosyl- (1----3)-O-alpha-L-rhamnopyranosyl-(1----4)-D-ribit-5-yl sodium phosphate] (27), which are structural elements of the capsular polysaccharides of Streptococcus pneumoniae types 6A and 6B ([----2)- -alpha-D-Galp-(1----3)-alpha-D-Glcp-(1----3)-alpha-L-Rhap- (1----X)-D-RibOH-(5-P----]n; 6A X = 3, 6B X = 4), respectively, have been synthesized. 2,4-Di-O-acetyl- 3-O-[2,4,6-tri-O-acetyl-3-O-(2,3,4,6-tetra-O-acetyl-alpha-D- galactopyranosyl)-alpha-D-glucopyranosyl]-alpha-L-rhamnopyranosyl trichloroacetimidate (13) was coupled with 5-O-allyloxycarbonyl-1,2,4-tri-O- benzyl-D-ribitol (10), using trimethylsilyl triflate as a promotor (----14), and deallyloxycarbonylation (----15) and conversion into the corresponding triethylammonium phosphonate then gave 16. Condensation of 16 with 4-methoxybenzyl 2,4-di-O-benzyl-3-O-[2,4,6-tri-O-benzyl-3-O-(3,4,6-tri-O-benzyl-alpha-D- galactopyranosyl)-alpha-D-glucopyranosyl]- alpha-L-rhamnopyranoside (22) followed by oxidation and deprotection afforded 25. 5-O-Allyl-1-O-allyloxycarbonyl-2,3-di-O-benzyl-D-ribitol (12) was coupled with 13, using trimethylsilyl triflate as a promoter, the resulting tetrasaccharide-alditol derivative 17 was deallyloxycarbonylated (----18), acetylated (----19), and deallylated (----20), and the product was converted into the triethylammonium phosphonate derivative 21. Condensation of 21 with 22 followed by oxidation and deprotection afforded 27.     

Oligosaccharide polyester and triterpenoid saponins from the roots of Polygala japonica

An oligosaccharide polyester, 1-O-(E)-p-coumaroyl-(3-O-benzoyl)-beta-D-fructofuranosyl-(2-->1)-[6-O-(E)-feruloyl-beta-D-glucopyranosyl-(1-->2)]-[6-O-acetyl-beta-D-glucopyranosyl-(1-->3)-(4-O-acetyl)-beta-D-glucopyranosyl-(1-->3)]-4-O-[4-O-alpha-L-rhamnopyranosyl-(E)-p-coumaroyl]-alpha-D-glucopyranoside (polygalajaponicose I), and four triterpenoid saponins, 3beta, 23, 27-trihydroxy-29-O-beta-D-glucopyranosyl-(1-->2)-beta-D-glucopyranosyl-olean-12-en-28-oic acid (polygalasaponin XLVII), 3-O-beta-D-glucopyranosyl presenegenin 28-O-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-fucopyranosyl ester (polygalasaponin XLVIII), 3-O-beta-D-glucopyranosyl presenegenin 28-O-beta-D-galactopyranosyl-(1-->5)-beta-D-apiofuranosyl-(1-->4)-beta-D-xylopyranosyl-(1-->4)-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-glucopyranosyl ester (polygalasaponin XLIX) and 2beta, 27-dihydroxy-3-O-beta-D-glucopyranosyl 11-oxo-olean-12-en-23, 28-dioic acid 28-O-beta-D-galactopyranosyl-(1-->5)-beta-D-apiofuranosyl-(1-->4)-beta-D-xylopyranosyl-(1-->4)-alpha-L-rhamnopyranosyl-(1-->2)-beta-D-fucopyranosyl ester (polygalasaponin L), in addition to five known compounds have been isolated from the roots of Polygala japonica.     

Bioactive sucrose esters from Bidens parviflora

An investigation on Bidens parviflora led to the isolation of three sucrose esters and a substituted truxillate. Their structures were elucidated as (6-O-(E)-p-coumaroyl)-beta-D-fructofuranosyl-(2-->1)-alpha-D-glucopyranoside, (6-O-(E)-p-coumaroyl)-beta-D-fructofuranosyl-(2-->1)-(6-O-(E)-p-coumaroyl)-alpha-D-glucopyranoside II, 6,6'-sucrose ester of (1alpha,2alpha,3beta,4beta)-3,4-bis(4-hydroxyphenyl)-1,2-cyclobutanedicarboxylic acid, dimethyl ester of (1alpha,2alpha,3alpha,4alpha)-2,4-bis(3,4-dihydroxyphenyl)-1,3-cyclobutanedicarboxylic acid on the basis of spectral and chemical evidence. These compounds were subjected to the following bioassays: the histamine release inhibition of rat mast cells induced by antigen-antibody reaction and the inhibitory activity of PGE(2) production by macrophages.     

Chemical characterization of the immunomodulating polysaccharide of Aloe vera L

The polysaccharide isolated by alcohol precipitation of Aloe vera mucilaginous gel was found to have a Man:Glc:Gal:GalA:Fuc:Ara:Xyl ratio of 120:9:6:3:2:2:1 with traces of Rha and GlcA. Linkage analysis of the endo-(1-->4)-beta-d-mannanase-treated sample yielded Manp-(1--> (approximately 26%), 4-Manp (approximately 53%), 2,4-Manp (approximately 3%), 3,4-Manp (approximately 1%), 4,6-Manp (approximately 1%), 4-Glcp (approximately 5%), 4-Xylp (approximately 1%), Xylp-(1--> (approximately 2%), Galp-(1--> (approximately 5%), and traces of 4,6-Galp and 3,6-Galp. Hydrolysis with strong acids produced a mixture of short oligosaccharides and an acid-resistant fraction containing greater relative fractions of Manp-(1-->, Araf-(1-->, Xylp-(1-->, and 4-Xylp than the bulk polysaccharide. NMR analysis of oligosaccharides generated by endo-(1-->4)-beta-D-mannanase and acid hydrolysis showed the presence of di-, tri-, and tetrasaccharides of 4-beta-Manp, beta-Glcp-(1-->4)-Man, beta-Glcp-(1-->4)-beta-Manp-(1-->4)-Man, and beta-Manp-(1-->4)-[alpha-Galp-(1-->6)]-Man, consistent with a backbone containing alternating -->4)-beta-Manp-(1--> and -->4)-beta-Glcp-(1--> residues in a approximately 15:1 ratio. Analysis of the sample treated sequentially with endo-(1-->4)-beta-d-mannanase and alpha-D-galactosidase showed that the majority of alpha-Galp-(1--> residues were linked to O-2, O-3, or O-6 of -->4)-beta-Manp-(1--> residues, with approximately 16 -->4)-beta-Manp-(1--> residues between side chains. Our data provide direct evidence of a previously proposed glucomannan backbone, but draw into question previously proposed side-chain structures.     

Recombinant (2-->3)-alpha-sialyltransferase immobilized on nickel-agarose for preparative synthesis of sialyl Lewis(x) and Lewis(a) precursor oligosaccharides

The specificity of recombinant (2-->3)-alpha-sialyltransferase (ST3Gal-III), expressed in baculovirus-infected insect cells, has been determined with various oligosaccharide acceptors and sugar-nucleotide donors using a fluorescence based assay. Recombinant ST3Gal-III tagged with a polyhistidine tail was immobilized on Ni(2+)-NTA-Agarose as an active enzyme for use in the synthesis of three sialylated oligosaccharides: (i) the divalent molecule [alpha-Neu5Ac-(2-->3)-D-Galp-(1-->4)-beta-D-GlcpNAc-O-CH(2)](2)-C-(CH(2)OBn)(2) (12); (ii) the dansylated derivative, alpha-Neu5Ac-(2-->3)-D-Galp-(1-->3)-beta-D-GlcpNAc-O-(CH(2))(6)-NH-dansyl and; (iii) the tetrasacharide alpha-Neu5Ac-(2-->3)-beta-D-Galp-(1-->4)-beta-D-GlcpNAc-(1-->2)-alpha-D-Manp-O-CH(3). Compound 12 was itself prepared from the divalent N-acetyllactosamine molecule built on pentaerythritol by a chemo-enzymatic route.     

Saponins and acylated saponins from Dizygotheca kerchoveana

Four new triterpenoid saponins were isolated from the leaves and stem of branches of Dizygotheca kerchoveana along with seven known ones. The new saponins were respectively characterized as 3-O-[beta-D-glucopyranosyl-(1-->3)]-[beta-D-glucopyranosyl-(1-->2)]-alpha-L-arabinopyranosyl echinocystic acid, 3-O-[beta-D-glucopyranosyl-(1-->3)]-[beta-D-glucopyranosyl-(1-->2)]-alpha-L-arabinopyranosyl echinocystic acid 28-O-[alpha-L-rhamnopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->6)-beta-D-glucopyranosyl] ester, 3-O-[beta-D-3-O-trans-p-coumaroyl-glucopyranosyl-(1-->3)]-[beta-D-glucopyranosyl-(1-->2)]-alpha-L-arabinopyranosyl echinocystic acid 28-O-[alpha-L-rhamnopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->6)-beta-D-glucopyranosyl] ester and 3-O-[beta-d-3-O-cis-p-coumaroyl-glucopyranosyl-(1-->3)]-[beta-D-glucopyranosyl-(1-->2)]-alpha-L-arabinopyranosyl echinocystic acid 28-O-[alpha-L-rhamnopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->6)-beta-D-glucopyranosyl] ester. Their structures were elucidated by 1D and 2D NMR experiments, FAB-MS as well as chemical means.     

Cloning and expression of levansucrase from Leuconostoc mesenteroides B-512 FMC in Escherichia coli

Leuconostoc mesenteroides B-512 FMC produces dextran and levan using sucrose. Because of the industrial importance of dextrans and oligosaccharides synthesized by dextransucrase (one of glycansucrases from L. mesenteroides), much is known about the dextransucrase, including expression and regulation of gene. However, no detailed report about levansucrase, another industrially important glycansucrase from L. mesenteroides, and its gene was available. In this paper, we report the first-time isolation and molecular characterization of a L. mesenteroides levansucrase gene (m1ft). The gene m1ft is composed of 1272-bp nucleotides and codes for a protein of 424 amino acid residues with calculated molecular mass of 47.1 kDa. The purified protein was estimated to be about 51.7 kDa including a His-tag based on SDS-PAGE. It showed an activity band at 103 kDa on a non-denaturing SDS-PAGE, indicating a dimeric form of the active M1FT. M1FT levan structure was confirmed by NMR and dot blot analysis with an anti-levan-antibody. M1FT converted 150 mM sucrose to levan (18%), 1-kestose (17%), nystose (11%) and 1,1,1-kestopentaose (7%) with the liberation of glucose. The M1FT enzyme produced erlose [O-alpha-D-glucopyranosyl-(1-->4)-O-alpha-D-glucopyranosyl-(1-->2)-beta-D-fructofuranoside] as an acceptor product with maltose. The optimum temperature and pH of this enzyme for levan formation were 30 degrees C and pH 6.2, respectively. M1FT levansucrase activity was completely abolished by 1 mM Hg2+ or Ag2+. The Km and Vmax values for levansucrase were calculated to be 26.6 mM and 126.6 micromol min-1 mg-1.     

Chemo-enzymatic synthesis of tetra-, penta-, and hexasaccharide fragments of the capsular polysaccharide of Streptococcus pneumoniae type 14

The chemo-enzymatic synthesis is described of beta-D-Glcp-(1-->6)-[beta-D-Galp-(1-->4)]-beta-D-GlcpNAc-(1-->3)-beta-D-Galp-(1-->O(CH(2))(6)NH(2) (1), beta-D-Glcp-(1-->6)-[beta-D-Galp-(1-->4)]-beta-D-GlcpNAc-(1-->3)-beta-D-Galp-(1-->4)-beta-D-Glcp-(1-->O(CH(2))(6)NH(2) (2), beta-D-Galp-(1-->4)-beta-D-GlcpNAc-(1-->3)-beta-D-Galp-(1-->4)-beta-D-Glcp-(1-->O(CH(2))(6)NH(2) (3), and beta-D-Galp-(1-->4)-beta-D-GlcpNAc-(1-->3)-beta-D-Galp-(1-->4)-beta-D-Glcp-(1-->6)-[beta-D-Galp-(1-->4)]-beta-D-GlcpNAc-(1-->O(CH(2))(6)NH(2) (4), representing fragments of the repeating unit of the Streptococcus pneumoniae serotype 14 capsular polysaccharide. Linear intermediate oligosaccharides 5-8 were synthesized via chemical synthesis, followed by enzymatic galactosylation using bovine milk beta-1,4-galactosyltransferase as a catalyst. The title oligosaccharides form suitable compounds for conjugation with carrier proteins, to be tested as potential vaccines in animal models.     

Synthesis, spectroscopical characterization and the biological activity in vitro of new platinum(II) complexes with imidazo[1,5-a]-1,3,5-triazine derivatives and dimethylsulfoxide

A series of platinum(II) complexes with 6,8-dimethylimidazo[1,5-a]-1,3,5-triazin-4(3H)-one (6,8-DiMe-4-O-IMT) (I) and 6,8-dimethyl-2-thioxo-2,3-dihydroimidazo[1,5-a]-1,3,5-triazin-4(1H)-one (6,8-DiMe-4-O-2-S-IMT) (II) of formula trans-[PtCl₂(dmso)(6,8-DiMe-4-O-IMT)] (1a) and trans-[PtCl₂(dmso)(6,8-DiMe-4-O-2-S-IMT)] (2a) have been prepared and characterized with ¹H, ¹³C, ¹⁵N, ¹⁹⁵Pt NMR and IR. Significant ¹⁵N NMR upfield coordination shifts (81-96 ppm) of N(7) atom indicate this nitrogen atom as a coordination site. The multinuclear NMR and IR spectra indicate the square planar geometry with N(7) bonded heterocycles, S-bonded dimethylsulfoxide and two trans chloride anions. The platinum(II) complexes were tested for their antiproliferative activity in vitro against the cells of four human cell lines: SW707 rectal adenocarcinoma, A549 non-small cell lung carcinoma, T47D breast cancer and HCV29T bladder cancer. The activity of (1a, 2a) was lower than that of cisplatin.     

Synthesis, characterization and DNA binding properties of oligopyridine-ruthenium(II)-amino acid conjugates

The DNA-binding properties of a number of ruthenium oligopyridine complexes with conjugated amino acids having the general formulae [Ru(terpy)(4-COY-4'-Mebpy)(X)](n)(+), X=NO (n=3), X=Cl (n=1) and NO(2) (n=1) and Y=AlaCONH(2) and TrpCONH(2) are reported. The new complexes were spectroscopically characterized and their DNA-binding properties were studied by means of circular dichroism (CD), (23)Na and (31)P NMR spectroscopy. The results show that the chlorido complexes interact by coordination to the DNA bases with the conjugated amino acid able to provide an additional interaction with the DNA helix. In addition, electrostatic interactions between all studied complexes and the DNA polyanion were observed. The nitro complexes were found to be insignificant, affecting only the (31)P NMR signal, probably due to changes in the hydration sphere of the DNA close to the phosphates.     

Structure elucidation by NMR spectroscopy of a new acetylated saponin from Centratherum anthelminticum

A new glycosylated triterpene has been isolated from the seeds of Centratherum anthelminticum, a medicinally important plant. The structural analysis of its acetylated derivative was performed by 1H, 13C NMR, 1H-1H COSY, HMQC, HMBC and DEPT spectroscopy. The saponin was shown to contain hederagenin and six sugar residues forming two glycosyl chains. The complete structure of the saponin was established as 3-O-[beta-D-glucopyranosyl-(1-->3)-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranosyl]-28-O-[beta-D-glucuronopyranosyl-(1-->4)-alpha-L-rhamnopyranosyl-(1-->3)-beta-D-glucopyranosyl]-hederagenin.     

Conformational studies of a novel cationic glycolipid,glyceroplasmalopsychosine, from bovine brain by NMR spectroscopy

A novel glycosphingolipid containing a long chain aldehyde conjugated to galactose and glycerol, Gro1(3)-O-CH((CH(2))(n)CH(3))-O-6Galbeta-sphingosine (glyceroplasmalopsychosine) has been studied by NMR spectroscopy (Hikita et al. J. Biol. Chem. 2001, 276, 23084-23091). We further report here on the conformation showing the galactose and the glycerol at the end of two parallel hydrophobic chains, i.e. the sphingosine and the fatty aldehyde. This is proposed based on the interproton distances derived from ROESY experiments and 3 J (H,H) coupling constants. The absence of any intraresidual NOEs between protons in the glycerol residue suggested that the C-C-2 and C-C-3 bonds in the glycerol may be rotating freely, supporting the proposed conformation in which the unique terminal glycerol is in an environment with a minimal steric hindrance. The present study proposes a conformation of glyceroplasmalopsychosine greatly different from the two conventional plasmalopsychosines possessing a fatty aldehyde chain oriented in an opposite direction to the sphingosine.     

Structural analysis of the alpha-D-glucan (EPS35-5) produced by the Lactobacillus reuteri strain 35-5 glucansucrase GTFA enzyme

The neutral exopolysaccharide EPS35-5 (reuteran) produced from sucrose by the glucansucrase GTFA enzyme from Lactobacillus reuteri 35-5 was found to be a (1-->4,1-->6)-alpha-D-glucan, with no repeating units present. Based on linkage analysis and 1D/2D 1H and 13C NMR spectroscopy of intact EPS35-5, as well as MS and NMR analysis of oligosaccharides obtained by partial acid hydrolysis and enzymatic hydrolysis, using pullulanase M1 (Klebsiella planticola), of EPS35-5, a composite model, that includes all identified structural elements, was formulated as follows: [Formula: see text].     

Triterpenoid saponins from Oreopanax guatemalensis

Seven oleanane-type saponins were isolated from the leaves and stems of Oreopanax guatemalensis, together with ten known saponins of lupane and oleanane types. The new saponins were respectively characterized as 3-O-alpha-L-arabinopyranosyl echinocystic acid 28-O-[alpha- L-rhamnopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->6)-beta-D-glucopyranosyl] ester, 3-O-beta-D-glucopyranosyl 3beta-hydroxy olean-11,13(18)-dien-28-oic acid 28-O-[alpha-L-rhamnopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->6)-beta- D-glucopyranosyl]ester, 3-O-[alpha-L-rhamnopyranosyl-(1-->2)-beta-D-glucopyranosyl]3beta-hydroxy olean-11,13(18)-dien-28-oic acid 28-O-[alpha-L-rhamnopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->6)-beta-D-glucopyranosyl] ester, 3-O-[alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranosyl]3beta, 23 dihydroxy olean-18-en-28-oic acid 28-O-[alpha-L-rhamnopyranosyl-(1-->4)-beta-D-6-O-acetyl glucopyranosyl-(1-->6)-beta-D-glucopyranosyl]ester, 3-O-[alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranosyl] hederagenin 28-O-[alpha-L-rhamnopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->6)-[beta-D-xylopyranosyl-(1-->2 )-]beta-D-glucopyranosyl]ester, 3-O-[alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranosyl]hederagenin 28-O-[alpha-L-rhamnopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->6)-[beta-D-glucopyranosyl-(1-->2)-]beta-D-glucopyranosyl] ester and 3-O-[alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranosyl] hederagenin 28-O-[alpha-L-rhamnopyranosyl-(1-->4)-beta-D-glucopyranosyl-(1-->6)-[alpha-L-arabinofuranosyl-(1-->2)]-beta-D-glucopyranosyl] ester. The structures were determined by spectral analyses. The NMR assignments were made by means of HOHAHA, 1H-1H COSY, HMQC, HMBC spectra and NOE difference studies.     

Synchronous fluorescence, UV-visible spectrophotometric, and voltammetric studies of the competitive interaction of bis(1,10-phenanthroline)copper(II) complex and neutral red with DNA

Constant wavelength synchronous fluorescence spectroscopy (CW-SFS), UV-visible absorption spectroscopy, and cyclic and differential pulse voltammetry were applied to investigate the competitive interaction of DNA with the bis(1,10-phenanthroline)copper(II) complex cation ([Cu(phen)(2)](2+)) and a fluorescence probe, neutral red dye (NR), in a tris-hydrogen chloride buffer (pH 7.4). The results show that both the [Cu(phen)(2)](2+)and the NR molecules can intercalate competitively into the DNA double-helix structure. The cyclic voltammetry method showed that both anodic and cathodic currents of [Cu(phen)(2)](2+) decreased on addition of the DNA and the intercalated [Cu(phen)(2)](2+)-DNA complex formed (beta = (4.14 +/- 0.24) x 10(3)). CW-SFS measurements were facilitated by the use of the three-way resolution of the CW-SFS for NR, [Cu(phen)(2)](2+), and NR-DNA. The important constant wavelength (CW) interval, Deltalambda, was shown to vary considerably when optimized (135, 58, and 98 nm for NR, NR-DNA, and [Cu(phen)(2)](2+), respectively). This approach clearly avoided the errors that otherwise would have arisen from the common assumption that Deltalambda is constant. Furthermore, a chemometrics approach, parallel factor analysis (PARAFAC), was applied to resolve the measured three-way CW-SFS data, and the results provided simultaneously the concentration information for the three reaction components, NR, [Cu(phen)(2)](2+), and NR-DNA, for the system at each equilibrium point. The PARAFAC analysis indicated that the intercalation of the [Cu(phen)(2)](2+) molecule into the DNA proceeds by exchanging with the NR probe and can be attributed to two parallel reactions. Comprehensive information was readily obtained; the replacement of the intercalated NR commenced immediately on introduction of [Cu(phen)(2)](2+), approximately 50% of NR was replaced by [Cu(phen)(2)](2+) at a concentration of 0.45 x 10(-5) mol L(-1), and nearly all of the NR was replaced at a [Cu(phen)(2)](2+) concentration of 2.50 x 10(-5) mol L(-1). This work has the potential to improve extraction of information from the fluorescence intercalator displacement (FID) assay.     

Heterodinuclear Cu(II)Zn(II) complexes: F NMR as a versatile tool to control the synthesis

The dinucleating ligands 2,6-bis[(bis(2-pyridylmethyl)amino)methyl]-4-methylphenol (H-BPMP) and 2,6-bis[(bis(2-pyridylmethyl)amino)methyl]-4-fluorophenol (H-BPFP) were used to synthesize heterodinuclear (μ-phenoxo)(μ-hydroxo) Cu(II)Zn(II) complexes. The labeled ligand with a fluorine atom allows the use of ¹⁹F NMR spectroscopy, which turned to be a rapid and powerful tool to tune the synthesis of the heterodinuclear paramagnetic complex [CuZnBPFP(μOH)](ClO₄)₂ and avoid mixing of complexes with statistical distribution. When applied to the non-fluorinated ligand, this experimental procedure leads to prepare and isolate easily the complex [CuZnBPMP(μOH)](ClO₄)₂. The X-ray structure is described.     

Enzymatic synthesis of 4-methylumbelliferyl (1-->3)-beta-D-glucooligosaccharides-new substrates for beta-1,3-1,4-D-glucanase

The transglycosylation reactions catalyzed by beta-1,3-D-glucanases (laminaranases) were used to synthesize a number of 4-methylumbelliferyl (MeUmb) (1-->3)-beta-D-gluco-oligosaccharides having the common structure [beta-D-Glcp-(1-->3)](n)-beta-D-Glcp-MeUmb, where n=1-5. The beta-1,3-D-glucanases used were purified from the culture liquid of Oerskovia sp. and from a homogenate of the marine mollusc Spisula sachalinensis. Laminaran and curdlan were used as (1-->3)-beta-D-glucan donor substrates, while MeUmb-beta-D-glucoside (MeUmbGlcp) was employed as a transglycosylation acceptor. Modification of [beta-D-Glcp-(1-->3)](2)-beta-D-Glcp-MeUmb (MeUmbG(3)) gives 4,6-O-benzylidene-D-glucopyranosyl or 4,6-O-ethylidene-D-glucopyranosyl groups at the non-reducing end of artificial oligosaccharides. The structures of all oligosaccharides obtained were solved by 1H and 13C NMR spectroscopy and electrospray tandem mass spectrometry. The synthetic oligosaccharides were shown to be substrates for a beta-1,3-1,4-D-glucanase from Rhodothermus marinus, which releases MeUmb from beta-di- and beta-triglucosides and from acetal-protected beta-triglucosides. When acting upon substrates with d.p.>3, the enzyme exhibits an endolytic activity, primarily cleaving off MeUmbGlcp and MeUmbG(2).     

Acetylated glucuronide triterpene bidesmosidic saponins from Symplocos glomerata

Nine new bidesmosidic 3-O-glucuronide oleanane triterpenoid saponins were isolated from the stem bark of Symplocos glomerata King along with two known saponins, salsoloside C and copteroside E, and two major lignans, (-)-pinoresinol and (-)-pinoresinol-4'-O-beta-D-glucopyranoside. The structures of the new saponins were established using one- and two-dimensional NMR spectroscopy and mass spectrometry as, 3-O-[beta-D-xylopyranosyl(1-->4)-[2-O-acetyl]-beta-D-glucuronopyranosyl]-28-O-[beta-D-glucopyranosyl]-oleanolic acid, 3-O-[beta-D-xylopyranosyl(1-->4)-[3-O-acetyl]-beta-D-glucuronopyranosyl]-28-O-[beta-D-glucopyranosyl]-oleanolic acid, 3-O-[beta-D-xylopyranosyl (1-->4)-[2,3-O-diacetyl]-beta-D-glucuronopyranosyl]-28-O-[beta-D-glucopyranosyl]-oleanolic acid, 3-O-[alpha-L-arabinopyranosyl(1-->4)-beta-D-glucuronopyranosyl]-28-O-[beta-D-glucopyranosyl]-oleanolic acid, 3-O-[alpha-L-arabinopyranosyl (1-->4)-[2-O-acetyl]-beta-D-glucuronopyranosyl]-28-O-[beta-D-glucopyranosyl]-oleanolic acid, 3-O-[[beta-D-xylopyranosyl (1-->2)]-[beta-D-xylopyranosyl (1-->4)]-[3-O-acetyl]-beta-D-glucuronopyranosyl]-28-O-[beta-D-glucopyranosyl]-oleanolic acid, 3-O-[[beta-D-glucopyranosyl (1-->2)]-[beta-D-xylopyranosyl (1-->4)]-[3-O-acetyl]-beta-D-glucuronopyranosyl]-28-O-[beta-D-glucopyranosyl]-oleanolic acid, 3-O-[[beta-D-glucopyranosyl (1-->2)]-[alpha-L-arabinofuranosyl (1-->4)]-[3-O-acetyl]-beta-D-glucuronopyranosyl]-28-O-[beta-D-glucopyranosyl]-oleanolic acid, and 3beta-O-[beta-D-xylopyranosyl(1-->4)-[2-O-acetyl]-beta-D-glucuronopyranosyl]-28-O-[beta-D-glucopyranosyl]-morolic acid. The EtOH and EtOAc extracts of the stem bark showed no cytotoxic activity. At a concentration of 370 microg/ml, the saponin mixture showed haemolytic activity and caused 50% haemolysis of a 10% suspension of sheep erythrocytes.     

A triterpenoidal saponin and sphingolipids from Pteleopsis hylodendron

From the stem bark of Pteleopsis hylodendron, a triterpenoidal saponin bellericagenin [B 3-O-[beta-D-glucopyranosyl-(1-->2)-alpha-D-glucopyranoside] (1) (Pteleopsoside)] and two sphingolipids, hylodendroside-I (2), and hylodendroside-II (3) were isolated, along with a synthetically known compound, [2alpha, 3beta, 23-triacetoxy-19alpha-hydroxyolean-12-en-28-oic acid (4)]. Other known compounds, friedelin (5), beta-carotene (6), lupeol (7), sitosterol (8), and stigmasterol (9), were also obtained. Their structures were deduced with the help of detailed spectroscopic studies.